Glycosyl transferases are a group of enzymes which are associated to the cell membrane and have been implicated to play a role for intercellular adhesion and tumor invasion. We are studying one of these enzymes, galactosyl transferase. We have purified this enzyme from both rat and human milk using two rapid chromatography steps, N-acetyl-glucosamine-sepharose and Alpha-lactalbumin-sepharose. This procedure enables the isolation of the rat milk enzyme which is highly labile and has low activity. This enzyme showed three polypeptide bands of MW 59K, 54K and 27K on SDS-polyacrylamide gel. The human milk enzyme on the other hand showed a single band of MW 54K on SDS-polyacrylamide gel. Antisera raised against the 54K form of rat milk galactosyl transferase immunoprecipitated the other forms suggesting that they are antigenically related. However, this antisera did not cross react with human milk enzyme.